Giardia lamblia is the most common disease-causing parasite in the United States responsible for an estimated 3 million cases a year. Besides causing problems in daycare centers, travelers, backpackers, and homosexuals, the parasite not uncommonly contaminates water supplies and has been responsible for massive epidemics. The organism lives and multiples in the small intestines and by unknown means causing diarrhea, abdominal pain, nausea and vomiting. Symptoms are frequently intermittent and long lasting. An environmentally resistant cyst form is passed in the feces and because large numbers of cysts are excreted and only a few can cause infection, giardiasis is a very common problem in the developed regions as well as undeveloped areas of the world. A major area of investigation is understanding how the parasite is able to survive in the body and how the body controls the infection. In animal models and likely in humans, some immunity to infection develops. Using mice made immune deficient in specific ways so that different parts of the immune system did not function, we were are to show that cellular immunity and, in particular, T cells were critical in the control of the infection. Previously, antibodies were felt to be most important in curing infection so these findings were unexpected. Experiments are underway to understand how T cells are able to control Giardia infections. These studies will lead to a clearer understanding of how immunity develops, to the characterization of the antigens important in provoking an immune response, and to the eventual development of a viable vaccine.One-way organisms evade the immune response of the host is to change its surface. In this way, the host is always responding to the surface of the parasite, which is either no longer present, or allowing the minority of parasites without the initial surface coat to be preferentially selected and thereby escaping the host?s protective immune responses. Previously, we showed that Giardia lamblia undergoes surface antigenic variation. The organism can use anywhere from 150-300 different surface antigens. Using unique clones whose surface antigens can be detected using specific antibodies, we showed that clones behave differently in mice that are genetically unable to respond to the parasite. Therefore, organisms that only differ in their surface antigens are either preferred or eliminated in a host that lacks developmental immunity. Therefore, individual surface antigens are biologically unique. Preliminary evidence last year suggested that X-irradiated gerbils showed different responses compared to SCID mice. These studies were reproduced and expanded using a panel of 8 different clone-Mab. Mice and irradiated gerbils were infected at the same time. These studies confirmed that certain VSPs were favored and differed amongst hosts. The evidence indicates that the importance of the surface antigens may be in allowing Giardia to adapt to different hosts and then be able to infect a greater number of different mammals. We also showed that changes in the surface antigen of the parasite in the host are caused by antibody response to the surface, and this is different from the immune responses that control the infection. Therefore, in the host two processes determine what surface antigens are found on the parasite in the intestine. The surface antigen must be compatible with the intestine of the host and not be recognized by previous or developing antibody responses that will not permit these VSPs to multiply in the intestine. Although we previously showed that only one VSP was found on the surface of the parasite, we were able to demonstrate that uncommonly two VSPs could be found on the surface. Dually expressing parasites were sorted by FACS twice and the VSP expression followed over time. Dually expressed parasites decreased over time so that at 36 hrs practically no parasites expressed two VSPs. This suggests dually expressing parasites are transient and likely represent parasites that are in the process of switching from one VSP to another. Using a panel of Mabs against Giardia. a number of cytoskeletal proteins were defined, characterized, and localized in the parasite. Ankyrin repeats were found to be particularly common in Giardia proteins and appear to be found in about 4% of all the proteins in Giardia, an unprecedented number for any organism. Analysis showed that ankyrin repeats in Giardia have evolved extensively into a large family of motifs that group to themselves. Ankyrin repeats are believed to bind one protein to another. In one instance, it was demonstrated that the ankyrin repeats appeared necessary to localize the protein correctly in the parasite.